茶多酚对大鼠肾缺血再灌注损伤保护作用实验研究

目的探讨茶多酚(Tea polyphenols TP)预处理对大鼠肾缺血再灌注损伤的保护作用及其机制。方法建立大鼠肾缺血再灌注损伤(renal ischemia reperfusion injury RIRI)模型。选用健康SD大鼠42只,随机分为3组:假手术组、单纯肾缺血再灌注损伤组,茶多酚预处理组。于肾缺血再灌注(renal ischemia reperfusion RIR)开始时刻、RIR后2h、RIR后24h分别检测血清肌酐(SCr),血清超氧化物岐化酶(SOD)、血清丙二醛(MDA)、肾组织SOD、肾组织MDA及观察肾组织的病理变化。结果茶多酚预处理组与假手术组相比较,RIRI组SCr水平升高(P<0.05),肾组织及血清中的SOD降低及MDA水平增加(P<0.05)。而茶多酚预处理组SCr、血清MDA和肾组织MDA均比单纯RIRI组低(P<0.05),血清SOD水平及肾组织SOD水平升高(P<0.05)。结论在肾脏缺血再灌注损伤过程中,茶多酚能起到对肾脏的保护作用。     

肢体急性动脉缺血再通术后再灌注损伤的防治

目的探讨急性肢体动脉缺血再通术后再灌注损伤的有效治疗方法。方法回顾性分析65例急性肢体动脉缺血再通术后再灌注损伤治疗患者的临床资料,上肢8例,下肢52例。结果65例急性肢体动脉缺血再通术后出现再灌注损伤38例。肢体急性动脉栓塞再通术后再灌注损伤程度较动脉硬化闭塞症继发急性血栓形成重,再通术后再灌注损伤程度与术前缺血时间及缺血程度成正比关系。动脉再通后30min出现再灌注损伤,24h达到高峰,72h后开始缓解,时间8～15d。肌筋膜切开3例,死亡2例,药物治愈33例。结论及时实施再通术是预防再灌注损伤的关键,再通术前后的有效治疗能够有效缓解再灌注损伤的发生和减轻再灌注损伤的症状。     

核转录因子NF-κB与肾脏缺血-再灌注损伤

核转录因子κB(NF-κB)普遍存在于真核生物细胞内,在免疫反应、炎症反应、细胞凋亡、缺血-再灌注损伤等病理生理改变过程中起着重要的调控作用。NF-κB在肾脏缺血-灌注损伤中的起着重要作用,本文就这方面的研究进展进行综述。     

西洋参皂苷对缺血-再灌注诱导乳鼠心肌细胞凋亡及钙浓度的影响

目的探讨西洋参皂苷(Panax quinquefolius saponin,PQS)对缺血-再灌注(ischemia/reperfusion,I/R)诱导乳鼠心肌细胞凋亡及钙浓度的影响。方法将原代培养的心肌细胞分为正常对照组(不做任何处理)、I/R组(心肌细胞经缺氧、无血清孵育2 h,再复氧、复血清培养24、48 h,以模拟心肌I/R损伤)和西洋参总皂苷干预组(PQS+I/R,细胞再灌注时分别加入5、10、20、40μg/ml PQS),采用台盼蓝染色法检测各组心肌细胞的活性;流式细胞术检测心肌细胞的凋亡率;激光扫描共聚焦显微镜检测心肌细胞内游离钙的变化。结果与I/R组比较,10、20、40μg/ml PQS+I/R组心肌细胞活性显著增加(P0.01),心肌细胞凋亡率显著降低(P0.05);20μg/ml的PQS可降低心肌细胞的钙负载。结论 PQS可显著减少I/R诱导的心肌细胞凋亡,并减轻心肌细胞钙超载的发生,这可能是PQS抑制I/R心肌细胞凋亡的机制之一。     

肝X受体激动剂对大鼠肝移植缺血再灌注损伤的保护作用

目的观察肝X受体(liver X receptor,LXR)激动剂GW3965预处理对大鼠肝移植缺血再灌注损伤(ischemiareperfution injury,IRI)的影响及其对肝功能的保护作用。方法将雄性SD大鼠随机分为2组,GW3965预处理组于供体大鼠尾静脉注射LXR激动剂GW3965,0.3 mg/kg;对照组于相同部位注射相同剂量的生理盐水。参照改进的Kamada两袖套法建立大鼠原位肝移植模型,分别于肝移植术后3、6、24 h,采用全自动生化分析仪检测血清谷丙转氨酶(alanine aminotransferase,ALT)含量,ELISA法检测血清中肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)含量,Western blot法测定移植肝脏组织中白细胞介素-1受体相关激酶-4(interleukin-1 receptor-associated kinase-4,IRAK-4)、干扰素调节因子3(interferon-regulator 3,IRF3)蛋白的表达,凝胶迁移试验(electrophoretic mobility shift assay,EMSA)测定核因子-kappa B(nuclear factor-κB,NF-κB)的相对活性度,并观察肝脏组织的病理学变化。结果与对照组比较,GW3965预处理组血清中ALT、TNF-α含量在各时间点均明显降低(P<0.05),肝脏组织中IRAK-4、IRF3蛋白的表达以及NF-κB相对活性度也明显降低(P<0.05);GW3965预处理组各时间点肝组织的损伤均较对照组明显减轻。结论 LXR激动剂GW3965可抑制TLR4信号通路中的IRAK-4、IRF3蛋白的表达水平和NF-κB的活化,从而减轻肝移植IRI,在肝脏移植IRI中起到保护作用。     

白藜芦醇对大鼠脑缺血再灌注氧化应激损伤的影响

目的探讨白藜芦醇(Resveratrol,Res)对大鼠脑缺血再灌注氧化应激损伤的影响。方法将SD大鼠随机分为假手术组(S组)、缺血再灌注组(I/R组,线栓法复制大鼠右侧大脑中动脉栓塞模型)、Res低剂量组(15 mg/kg,I/R+R1组)和Res高剂量组(30 mg/kg,I/R+R2组),于缺血2 h再灌注24 h进行神经功能缺损评分;化学比色法测定大鼠血清和脑组织中丙二醛(Malondialdehyde,MDA)含量及超氧化物歧化酶(Superoxide dismutase,SOD)活性;TTC法测定脑梗死体积;干湿重法测定脑含水量,HE染色观察脑组织的病理改变。结果与I/R组相比,Res能改善大鼠脑缺血再灌注损伤后的神经功能缺失(P<0.01),降低血清及脑组织中MDA的含量(P<0.01),提高SOD活性(P<0.01),缩小脑梗死体积(P<0.01),降低损伤侧脑含水量(P<0.01),改善脑组织的病理变化,且呈剂量依赖性。结论 Res对大鼠局灶性脑缺血再灌注氧化应激损伤具有良好的保护作用,其机制可能与清除自由基,减轻氧化性损伤有关。     

依达拉奉对大鼠脑缺血再灌注损伤的保护

目的探讨依达拉奉对大鼠脑缺血再灌注损伤的影响和作用机制。方法制备SD大鼠MCAO模型,梗死2h后再灌注,并分为假手术组、生理盐水(NS)组、依达拉奉低剂量组及依达拉奉高剂量组,再灌注24h后检测各组脑组织SOD、MDA浓度。结果与假手术组相比,NS组SOD活性降低、MDA含量增加,有显著性差异;与NS组相比,依达拉奉低剂量与高剂量组SOD活性增高、MDA含量降低,且低剂量组与高剂量组相比有显著性差异。结论依达拉奉对脑缺血再灌注损伤有保护作用。     

腺苷对缺血再灌注后心肌的保护作用

目的研究腺苷对大鼠心肌细胞凋亡及核因子κB(NF-κB)表达的影响。方法制备对照组(C组)、缺血再灌注损伤组(I/R组)和缺血再灌注前腺苷治疗组(AD组)的大鼠模型。电镜、光镜下观察心肌结构变化,采用原位缺口末端标记法(TUNEL)检测心肌细胞凋亡,免疫组织化学方法检测心肌组织NF-κB表达。结果AD组大鼠心肌细胞凋亡数(2 645.0±326.0)及心肌组织中NF-κB的表达(32.21%±17.91%)明显低于I/R组(5 113.0±503.7和60.30%±10.36%),明显高于对照组(67.7±51.3和11.98%±3.65%)。结论腺苷具有明显降低大鼠缺血再灌注后心肌细胞凋亡的作用,可能与腺苷减轻缺血再灌注心肌组织过度表达NF-κB有关。     

热量限制对小鼠心肌缺血/再灌注损伤的影响及其作用机制

目的 探讨热量限制(caloric restriction,CR)对小鼠心肌缺血/再灌注损伤(myocardial ischemia/reperfusion injury,MI/RI)的影响及其作用机制。方法 将C57小鼠随机分为正常饮食组(AL组,自由摄食)和CR组(饮食量每2周递减10%),持续8周,监测体质量变化。每组再分为假手术组和MI/RI组,共4组,即AL+Sham组、AL+I/R组和CR+Sham组、CR+I/R组,其中MI/RI组小鼠结扎冠状动脉左前降支30 min后,再灌注24 h;Sham组只穿线不结扎。伊文思兰/TTC染色法检测小鼠心肌缺血和梗死面积;HE染色法对心肌组织进行病理学观察;相应试剂盒检测心肌组织中乳酸脱氢酶(lactate dehydrogenase,LDH)、超氧化物歧化酶(superoxide dismutase,SOD)活性及肌酸激酶同工酶(creatine kinase-MB,CK-MB)、丙二醛(malondialdehvde,MDA)含量;ELISA法检测血清中IL-1β及IL-18含量;Western blot法检测心肌组织中细胞焦亡相...     

Canopy2对大鼠心肌缺血再灌注损伤心肌细胞凋亡的影响

目的 探讨 Canopy2(Canopy FGF signaling regulator 2,CNPY2)对大鼠心肌缺血再灌注损伤(myocardial ischemia-reperfusion injury,MIRI)心肌细胞凋亡的影响.方法 H2O2分别作用于感染腺病毒Ade-hCNPY2-Flag和Ade-NC-...     

Achyranthes bidentata Polypeptides Reduces Oxidative Stress and Exerts Protective Effects against Myocardial Ischemic/Reperfusion Injury in Rats

Achyranthes bidentata, a Chinese medicinal herb, is reported to be neuroprotective. However, its role in cardioprotection remains largely unknown. Our present study aimed to investigate the effects of Achyranthes bidentata polypeptides (ABPP) preconditioning on myocardial ischemia/reperfusion (MI/R) injury and to test the possible mechanisms. Rats were treated with ABPP (10 mg/kg/d, i.p.) or saline once daily for one week. Afterward, all the animals were subjected to 30 min of myocardial ischemia followed by 4 h of reperfusion. ABPP preconditioning for one week significantly improved cardiac function following MI/R. Meanwhile, ABPP reduced infarct size, plasma creatine kinase (CK)/lactate dehydrogenase (LDH) activities and myocardial apoptosis at the end of reperfusion in rat hearts. Moreover, ABPP preconditioning significantly inhibited superoxide generation, gp91^phox expression, malonaldialdehyde formation and enhanced superoxide dismutase activity in I/R hearts. Furthermore, ABPP treatment inhibited PTEN expression and increased Akt phosphorylation in I/R rat heart. PI3K inhibitor wortmannin blocked Akt activation, and abolished ABPP-stimulated anti-oxidant effect and cardioprotection. Our study demonstrated for the first time that ABPP reduces oxidative stress and exerts cardioprotection against MI/R injury in rats. Inhibition of PTEN and activation of Akt may contribute to the anti-oxidant capacity and cardioprotection of ABPP.     

Unilateral Acute Renal Ischemia-Reperfusion Injury Induces Cardiac Dysfunction through Intracellular Calcium Mishandling

Background: Acute renal failure (ARF) following renal ischemia-reperfusion (I/R) injury is considered a relevant risk factor for cardiac damage, but the underlying mechanisms, particularly those triggered at cardiomyocyte level, are unknown. Methods: We examined intracellular Ca²⁺ dynamics in adult ventricular cardiomyocytes isolated from C57BL/6 mice 7 or 15 days following unilateral renal I/R. Results: After 7 days of I/R, the cell contraction was significantly lower in cardiomyocytes compared to sham-treated mice. It was accompanied by a significant decrease in both systolic Ca²⁺ transients and sarco/endoplasmic reticulum Ca²⁺-ATPase (SERCA_2a) activity measured as Ca²⁺ transients decay. Moreover, the incidence of pro-arrhythmic events, measured as the number of Ca²⁺ sparks, waves or automatic Ca²⁺ transients, was greater in cardiomyocytes from mice 7 days after I/R than from sham-treated mice. Ca²⁺ mishandling related to systolic Ca²⁺ transients and contraction were recovered to sham values 15 days after I/R, but Ca²⁺ sparks frequency and arrhythmic events remained elevated. Conclusions: Renal I/R injury causes a cardiomyocyte Ca²⁺ cycle dysfunction at medium (contraction-relaxation dysfunction) and long term (Ca²⁺ leak), after 7 and 15 days of renal reperfusion, respectively.     

Bone Morphogenetic Protein-7 Ameliorates Cerebral Ischemia and Reperfusion Injury via Inhibiting Oxidative Stress and Neuronal Apoptosis

Previous studies have indicated that bone morphogenetic protein-7 (BMP-7) is neuroprotective against cerebral ischemia/reperfusion (IR) injury. The present study was undertaken to determine the molecular mechanisms involved in this effect. Adult male Wistar rats were subjected to 2 h of transient middle cerebral artery occlusion (MCAO), followed by 24 h of reperfusion. BMP-7 (10⁻⁴ g/kg) or vehicle was infused into rats at the onset of reperfusion via the tail vein. Neurological deficits, infarct volume, histopathological changes, oxidative stress-related biochemical parameters, neuronal apoptosis, and apoptosis-related proteins were assessed. BMP-7 significantly improved neurological and histological deficits, reduced the infarct volume, and decreased apoptotic cells after cerebral ischemia. BMP-7 also markedly enhanced the activities of antioxidant enzymes superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX), and reduced the level of malondialdehyde (MDA) in IR rats. In addition, Western blot analysis indicated that BMP-7 prevented cytochrome c release, inhibited activation of caspase-3, caspase-9 and caspase-8. Our data suggested that BMP-7 has protective effects against cerebral IR injury in rats, and the neuroprotective effects may be attributed to attenuating oxidative stress and inhibiting neuronal apoptosis.     

Effect of Ligusticum wallichii Aqueous Extract on Oxidative Injury and Immunity Activity in Myocardial Ischemic Reperfusion Rats

We investigated the efficacy of Ligusticum wallichi aqueous extract (LWE) for myocardial protection against ischemia-reperfusion injury. Rats were fed for five weeks with either a control diet (sham and ischemia reperfusion (IR) model control groups) or a diet mixed with 0.2%, 0.4% or 0.6% Ligusticum wallichi extract. At the end of the five week period, hearts were excised and subjected to global ischemia for 30 min followed by reperfusion for 2 h. The hearts were compared for indices of oxidative stress and immunity activities. Administration of Ligusticum wallichi extract significantly decreased serum TNF-α, IL-6, IL-8, NO, MIP-1α, CRP and myocardium MDA levels, and serum CK, LDH and AST activities, and increased myocardium Na(+)-K(+)-ATPase, Ca(2+)-Mg(2+)-ATPase, NOS, SOD, CAT, GSH-Px and TAOC activities. The results indicate that Ligusticum wallichii extract treatment can enhance myocardial antioxidant status and improve the immunity profile in ischemic-reperfusion rats.     

Hypothermia Alleviates Reductive Stress,a Root Cause of Ischemia Reperfusion Injury

Ischemia reperfusion injury is common in transplantation. Previous studies have shown that cooling can protect against hypoxic injury. To date, the protective effects of hypothermia have been largely associated with metabolic suppression. Since kidney transplantation is one of the most common organ transplant surgeries, we used human-derived renal proximal tubular cells (HKC8 cell line) as a model of normal renal cells. We performed a temperature titration curve from 37 °C to 22 °C and evaluated cellular respiration and molecular mechanisms that can counteract the build-up of reducing equivalents in hypoxic conditions. We show that the protective effects of hypothermia are likely to stem both from metabolic suppression (inhibitory component) and augmentation of stress tolerance (activating component), with the highest overlap between activating and suppressing mechanisms emerging in the window of mild hypothermia (32 °C). Hypothermia decreased hypoxia-induced rise in the extracellular lactate:pyruvate ratio, increased ATP/ADP ratio and mitochondrial content, normalized lipid content, and improved the recovery of respiration after anoxia. Importantly, it was observed that in contrast to mild hypothermia, moderate and deep hypothermia interfere with HIF1 (hypoxia inducible factor 1)-dependent HRE (hypoxia response element) induction in hypoxia. This work also demonstrates that hypothermia alleviates reductive stress, a conceptually novel and largely overlooked phenomenon at the root of ischemia reperfusion injury.     

Leonurine Reduces Oxidative Stress and Provides Neuroprotection against Ischemic Injury via Modulating Oxidative and NO/NOS Pathway

Leonurine (Leo) has been found to have neuroprotective effects against cerebral ischemic injury. However, the exact molecular mechanism underlying its neuroprotective ability remains unclear. The aim of the present study was to investigate whether Leo could provide protection through the nitric oxide (NO)/nitric oxide synthase (NOS) pathway. We firstly explored the effects of NO/NOS signaling on oxidative stress and apoptosis in in vivo and in vitro models of cerebral ischemia. Further, we evaluated the protective effects of Leo against oxygen and glucose deprivation (OGD)-induced oxidative stress and apoptosis in PC12 cells. We found that the rats showed anxiety-like behavior, and the morphology and number of neurons were changed in a model of photochemically induced cerebral ischemia. Both in vivo and in vitro results show that the activity of superoxide dismutase (SOD) and glutathione (GSH) contents were decreased after ischemia, and reactive oxygen species (ROS) and malondialdehyde (MDA) levels were increased, indicating that cerebral ischemia induced oxidative stress and neuronal damage. Moreover, the contents of NO, total NOS, constitutive NOS (cNOS) and inducible NOS (iNOS) were increased after ischemia in rat and PC12 cells. Treatment with L-nitroarginine methyl ester (L-NAME), a nonselective NOS inhibitor, could reverse the change in NO/NOS expression and abolish these detrimental effects of ischemia. Leo treatment decreased ROS and MDA levels and increased the activity of SOD and GSH contents in PC12 cells exposed to OGD. Furthermore, Leo reduced NO/NOS production and cell apoptosis, decreased Bax expression and increased Bcl-2 levels in OGD-treated PC12 cells. All the data suggest that Leo protected against oxidative stress and neuronal apoptosis in cerebral ischemia by inhibiting the NO/NOS system. Our findings indicate that Leo could be a potential agent for the intervention of ischemic stroke and highlighted the NO/NOS-mediated oxidative stress signaling.     

Chrysin Protects against Focal Cerebral Ischemia/Reperfusion Injury in Mice through Attenuation of Oxidative Stress and Inflammation

Inflammation and oxidative stress play an important part in the pathogenesis of focal cerebral ischemia/reperfusion (I/R) injury, resulting in neuronal death. The signaling pathways involved and the underlying mechanisms of these events are not fully understood. Chrysin, which is a naturally occurring flavonoid, exhibits various biological activities. In this study, we investigated the neuroprotective properties of chrysin in a mouse model of middle cerebral artery occlusion (MCAO). To this end, male C57/BL6 mice were pretreated with chrysin once a day for seven days and were then subjected to 1 h of middle cerebral artery occlusion followed by reperfusion for 24 h. Our data show that chrysin successfully decreased neurological deficit scores and infarct volumes, compared with the vehicle group. The increases in glial cell numbers and proinflammatory cytokine secretion usually caused by ischemia/reperfusion were significantly ameliorated by chrysin pretreatment. Moreover, chrysin also inhibited the MCAO-induced up-regulation of nuclear factor-kappa B (NF-κB), cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS), compared with the vehicle. These results suggest that chrysin could be a potential prophylactic agent for cerebral ischemia/reperfusion (I/R) injury mediated by its anti-inflammatory and anti-oxidative effects.     

Autophagy Mediates Escherichia Coli-Induced Cellular Inflammatory Injury by Regulating Calcium Mobilization,Mitochondrial Dysfunction,and Endoplasmic Reticulum Stress

Bovine endometritis is a reproductive disorder that is induced by mucus or purulent inflammation of the uterine mucosa. However, the intracellular control chain during inflammatory injury remains unclear. In the present study, we found that E. coli activated the inflammatory response through the assembly of the NLRP3 inflammasome and activation of the NF-κB p65 subunit in primary bovine endometrial epithelial cells (bEECs). Infection with E. coli also led to an abnormal increase in cytoplasmic calcium and mitochondrial dysfunction. Additionally, live-cell imaging of calcium reporters indicated that the increase in cytosolic calcium mainly was caused by the release of Ca²⁺ ions stored in the ER and mitochondria, which was independent of extracellular calcium. Cytoplasmic calcium regulates mitochondrial respiratory chain transmission, DNA replication, and biogenesis. Pretreatment with NAC, BAPTA-AM, or 2-APB reduced the expression of IL-1β and IL-18. Moreover, ERS was involved in the regulation of bovine endometritis and cytosolic calcium was an important factor for regulating ERS in E. coli-induced inflammation. Finally, activation of autophagy inhibited the release of IL-1β and IL-18, cytochrome c, ATP, ERS-related proteins, and cytoplasmic calcium. Collectively, our findings demonstrate that autophagy mediated E. coli-induced cellular inflammatory injury by regulating cytoplasmic calcium, mitochondrial dysfunction, and ERS.     

Insulin and α-Tocopherol Enhance the Protective Effect of Each Other on Brain Cortical Neurons under Oxidative Stress Conditions and in Rat Two-Vessel Forebrain Ischemia/Reperfusion Injury

Clinical trials show that insulin administered intranasally is a promising drug to treat neurodegenerative diseases, but at high doses its use may result in cerebral insulin resistance. Identifying compounds which could enhance the protective effects of insulin, may be helpful to reduce its effective dose. Our aim was thus to study the efficiency of combined use of insulin and α-tocopherol (α-T) to increase the viability of cultured cortical neurons under oxidative stress conditions and to normalize the metabolic disturbances caused by free radical reaction activation in brain cortex of rats with two-vessel forebrain ischemia/reperfusion injury. Immunoblotting, flow cytometry, colorimetric, and fluorometric techniques were used. α-T enhanced the protective and antioxidative effects of insulin on neurons in oxidative stress, their effects were additive. At the late stages of oxidative stress, the combined action of insulin and α-T increased Akt-kinase activity, inactivated GSK-3beta and normalized ERK1/2 activity in cortical neurons, it was more effective than either drug action. In the brain cortex, ischemia/reperfusion increased the lipid peroxidation product content and caused Na⁺,K⁺-ATPase oxidative inactivation. Co-administration of insulin (intranasally, 0.25 IU/rat) and α-T (orally, 50 mg/kg) led to a more pronounced normalization of the levels of Schiff bases, conjugated dienes and trienes and Na⁺,K⁺-ATPase activity than administration of each drug alone. Thus, α-T enhances the protective effects of insulin on cultured cortical neurons in oxidative stress and in the brain cortex of rats with cerebral ischemia/reperfusion injury.